This thesis presents the first detailed study of the torsional optokinetic nystagmus (tOKN) response in the normal population and in patients with oculomotor disorders. The effects on the tOKN response of: (i) stimulus velocity, (ii) stimulus area, and (iii) aging, were investigated in the normal population. The tOKN response was also evaluated in patients with long standing oculomotor disorders, namely strabismus and infantile nystagmus. Torsional OKN responses were recorded using infrared video-oculography and were elicited with volunteers fixating the centre of a large-field rotating sinusoidal grating pattern. Torsional OKN responses were present in all normal young volunteers (n=20) to stimuli rotated in clockwise and anticlockwise directions, and a linear relationship was observed between log stimulus velocity and tOKN slow phase velocity. Torsional OKN also showed brisk responses to peripheral field stimulation in the same subjects. The first report of a significant increase with age in the proportion of absent tOKN responses is also described in normal subjects aged between 19-72 years (n=30). The tOKN response was investigated for the first time in strabismic patients (n=16), comparing horizontal and vertical OKN responses, and also in patients with infantile nystagmus (n=16). OKN responses from strabismus patients demonstrated consistent asymmetry in horizontal and vertical directions. However, a significantly higher incidence of absent tOKN responses in both intorsion and extorsion directions were observed in comparison to controls. Torsional OKN was present in 3 of 16 patients with infantile nystagmus. Torsional OKN is a well developed reflex in the normal population with the capacity to respond in proportion to stimulus velocity and area of stimulation. However, the tOKN response is dramatically affected by (i) the effects of aging, (ii) by the interruption of binocular visual development in patients with strabismus, and (iii) by the presence of infantile nystagmus.

Background: Amblyopia, ‘lazy’ eye is a unilateral or bilateral reduction in vision for which no organic cause is present by physical examination of the eye with a prevalence of approximately 3.5% of the childhood population. It is commonly associated with a strabismus, refractive error or both. The most common form of treatment is conventional occlusion (daily patching the good eye). Clinical studies have attempted to investigate the optimal treatment of the disease and investigate compliance, however an evidence-base for treatment is still incomplete. Methods: The study included (i) a retrospective study of 322 amblyopic children to assess current visual outcomes in comparison to clinical effort and costs; (ii) A randomised control trial (n=52) comparing prescribed treatments of 0-hours, 3-hours and 6-hours patching per day in which compliance was electronically recorded; (iii) interviews of 25 families to explore reasons behind poor compliance; and (iv) a pilot study of educational material to improve compliance. Results: Current outcomes of amblyopia treatment are mediocre at considerable financial and time-costs. The RCT revealed poor compliance in both patching groups (3-hours and 6-hours) leading to visual improvements that were not significantly better than no patching. However, there was a clear dose-response between visual improvement and effective hours patched (p=0.00013). The interviews demonstrated emotional distress in families, lack of social acceptance, and confusion about amblyopia, its treatment and the role of professionals. Early findings indicate that an educational intervention could reduce the number of poor compliers. Conclusion: Poor compliance leads to poor visual outcomes of occlusion treatment for amblyopia. However, objective monitoring of patching demonstrates that occlusion therapy is effective. An educational intervention could address some of the problems associated with poor compliance such as poor parental understanding, providing feedback of visual improvement to the family and strategies for implementing patching as a normal routine.

Background: Nystagmus can be a manifestation of ocular or systemic disorders. However, it may represent a separate disease entity by itself as in idiopathic infantile nystagmus (IIN). In 2004, Kerrison et al. localised the gene causing X-linked IIN to Xq26-27 (NYS1); however, the gene/genes causing IIN had not been identified. Aims and Objectives: The aims of this study were threefold. 1. To ascertain families and singletons (sporadic subjects) with IIN. 2. To further refine the locus NYS1 and to identify the gene causing X-linked IIN 3. To describe and compare the phenotype of subjects with IIN Methods: 39 families and 78 singletons with nystagmus were recruited and phenotyped. Genotyping with microsatellite markers were performed in the X-linked families to refine the genetic interval at Xq26-27. Gene sequencing was carried out by our collaborators (not by the author) at the Sanger Institute. The clinical features and eye movement recordings of 90 subjects with mutations in the FRMD7 gene were compared to 48 subjects with IIN not associated with mutations in this gene (non-FRMD7 group). Results: I: 149 familial subjects and 78 sporadic subjects with IIN were phenotyped. 121 subjects from 30 families were diagnosed to have X-linked IIN while 28 subjects from 9 families had other diagnosis such as albinism and aniridia. II: Genetic mapping in 16 families with X-linked IIN, refined the critical interval at Locus NYS1 (Xq26-17) to a 9mB region between markers DXS8072 and DXS8094 which contained about 80 genes. High throughput DNA sequencing was carried out at the Sanger Institute which led to the discovery of FRMD7, mutations in which is associated with X-linked IIN. III: The median visual acuity in subjects with a FRMD7 mutation was log MAR 0.301. The number of subjects with good stereopsis (Lang positive) was higher in the FRMD7 group (93.4%) compared to subjects in the non- FRMD7 group (78.4%). None of the subjects in the FRMD7 group had severe (>15˚) anomalous head posture (AHP) while 27% of subjects in the non-FRMD7 group had AHP more than 15˚. 52.17% of obligate female carriers of a FRMD7 mutation were clinically affected. Discussion: This study identified the first gene causing idiopathic infantile nystagmus. The phenotypic characteristics of these subjects will help in clinically identifying subjects with IIN due to mutations in FRMD7. In addition it has generated a stage for further research into the mechanisms behind ocular motor control.

This study investigated the feasibility of assessing fundus oxygen perfusion in the human eye by adapting a spectral confocal scanning laser ophthalmoscope (cSLO). The Aberdeen prototype cSLO was adapted to use two wavelengths simultaneously performing a spectral analysis of the status of the retina tissue to give the ability of imaging the perfusion in the human eye. In order to image retinal perfusion a new combination of two wavelengths (red and infrared) was chosen to produce a linear relationship between oxygen saturation and optical density ratio of blood according to the Beer & Lambert law. The technique was evaluated using measurements made on a model eye, fourteen volunteers' normal eyes and seven eyes containing diabetic retinopathy. The reproducibility of the measurements of reflected light for the eye using the Aberdeen SLO was investigated. A model eye was designed and varieties of different elements of the SLO using the model were studied. It also was necessary to process SLO images to give uniform data improving the reproducibility of the measurements. So, an algorithm for image correction was developed to minimize the intensity variation across the image. The relative oxygen levels of retinal blood vessels (artery and vein) were successfully determined using this method. Retinal perfusion imaging (RPI) is a new technique that showed promise in the diagnosis of the non-perfused parts of the retina as well as normal regions using a pseudo-colour map. It has been shown RPI using the cSLO has advantage over fluorescein angiogram in that it can image relative oxygen saturation levels of the retina.

Mutations in PRPF31 have been implicated in autosomal dominant retinitis pigmentosa (adRP) for over a decade, yet the molecular basis underlying the observed phenotypic non-penetrance in families remains unknown. In the population, there are differentially expressed alleles of PRPF31: both high-expressivity and a low-expressivity alleles. Inheritance of a higher-expressivity allele alongside a mutant allele provides protection against the clinical manifestations of the disease. The aim of the research was to investigate phenotypic non-penetrance associated with PRPF31 mutations, through the understanding of transcriptional regulation of PRPF31 in both health and disease. Firstly, it was investigated whether the non-penetrant effect was caused by a cis- or trans-acting factor. Microsatellite data was used to show that inheritance of the wildtype chromosome 19q13 determined affection status of individuals harbouring a mutation in PRPF31. It was, therefore, concluded that a cis-acting element controls the non-penetrant phenotype. A family affected by adRP with a history of non-penetrance was identified and a large deletion (112kb) in the 19q13.4 region encompassing PRPF31 was fully characterized. Importantly, the deletion encompassed all introns of PRPF31 and also a large region upstream, therefore including putative regulatory elements of the gene. Dual-luciferase reporter assay was performed to define the core promoter of PRPF31 and also the core promoter of TFPT, a gene lying in a head-to-head arrangement with PRPF31, with partially shared exon 1. Core promoters were defined for both genes. A patient with a history of isolated adRP was identified, in whom a single base pair deletion in the PRPF31 core promoter was present. Functional studies were performed, demonstrating that the mutant allele had a serious adverse affect on transcription of PRPF31- this patient represents the first report of functional haploinsufficiency as a disease mechanism in adRP. Comparison of the PRPF31 core promoter sequence in an asymptomatic and a symptomatic individual lead to the identification of one polymorphism that significantly affected transcriptional activation of the gene. The duplication of a 38bp repeat element increased transcription, and it was found that this element was statistically under-represented in a cohort of symptomatic individuals compared to the general population. This represents the first identification of a molecular factor controlling non-penetrance in PRPF31-associated adRP. Finally, the evolution of the PRPF31 and TFPT core promoters was studied. The conservation of the region was analysed and regions homologous to the human core promoter fragments were sought in monkey, dog and mouse. Dual-reporter luciferase assay was performed to characterise the promoter elements in these three species, and demonstrated that conservation of a gene was not always accompanied by conservation of regulatory elements.

Pigment dispersion syndrome (PDS) is an ocular condition predisposing to glaucomatous optic neuropathy in patients at a relatively young age. Concavity of the iris is considered to be important in the pathogenesis of PDS, however, it is also appears to be a feature of non-PDS eyes, particularly in young myopes. Much of the current understanding of anterior segment anatomy is derived from studies using ultrasound biomicroscopy, a relatively invasive imaging modality that involves direct ocular contact. Anterior-segment optical coherence tomography (AS-OCT) allows imaging of the anterior segment with the patient in the upright position without the need for contact with the ocular surface. AS-OCT may allow a more physiological assessment of anterior segment anatomy as well as being better suited to paediatric subjects. AS-OCT was used to conduct a case-control study of anterior segment anatomy in PDS subjects and age-, sex- and refraction- matched controls to determine which features of anterior segment anatomy best discriminated between the 2 groups. In addition AS-OCT was used to assess anterior segment anatomy, with particular emphasis on iris curvature, in a cohort of 10-12 year old school children and explore correlations with ocular biometry and parameters reflecting corneal biomechanical properties. Longitudinal data was collected through re-visiting the cohort 2 years later. Chromosomal susceptibility loci for PDS have been described, although no causative gene has been identified. Two approaches were used to identify novel disease susceptibility loci: 1) linkage analysis was used in a 3-generation family segregating for PDS/pigmentary glaucoma, and, 2) DNA from a large cohort of unrelated PDS probands was collected and sent for genotyping with a view to conducting a pilot genome-wide association study. Finally a candidate gene, GPNMB, the human homologue of a causative gene in a mouse model of pigmentary glaucoma was sequenced in a panel of 96 unrelated PDS/pigmentary glaucoma subjects.

No description available.

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The search for novel biomarkers and therapeutic targets is a continuous process in the fight against disease. In this project, target identification and validation approaches were used towards the discovery of protein targets related to distinct, but equally relevant diseases. The first approach involved the use of proteomics to identify protein targets in infertility. An affinity-enrichment proteomic method was developed and successfully applied to the identification of protein targets of a clinically utilised compound, which is known to cause reversible, dose-dependent male infertility in certain mouse strains. The second approach demonstrated the versatility of proteomics in an attempt to answer a completely different question. In this study, it was applied to the discovery of protein targets of chemoresistance in ovarian cancer, using human ovarian cancer cell lines and tissue biopsies. Once again, this method was successful in identifying some very promising un-regulated protein targets and pathways involved in cancer resistance. Finally, a more assay development orientated approach aimed for the functional characterisation of Hsp90 targeted compounds. The combined use of a native gel binding assay and an Hsp90 ATPase assay proved to be a convenient and robust method to characterise Hsp90 inhibitors and will aid the development of Hsp90 targeted anti-cancer drugs. In summary, the work undertaken confirmed the recognised advantages of proteomics in the comparative study of un-regulated proteins connected to disease. It equally showed how two distinct techniques could be used in synergy to accomplish more valuable answers, in the screening of inhibitors of a known protein target.

No description available.