ABSTRACT Purpose: The primary aim of the study (i.e. Chapter 3) was to compare the magnitude of retinal vascular reactivity in arterioles of varying diameter in healthy, young subjects. The secondary aims were to determine: a) if there are any order effects in terms of provoking vasoconstriction or vasodilation first; and b) the repeatability of the vascular reactivity measurements. An additional aim (i.e. Chapter 4) was to determine the effect of healthy aging on the relationship between retinal vascular reactivity and vessel diameter. Method: The sample comprised 10 healthy, young subjects (mean age 26.5 years, SD 4.04) and 7 healthy, older subjects (mean age 55.43 years, SD 5.41). Each subject from the young age group attended for three sessions. The first session was used to determine eligibility and select hemodynamic measurement sites. At sessions 2 and 3, O2 and CO2 were sequentially administered to the subjects using a face mask and sequential re-breathing circuit (to maintain standardized hyperoxia and hypercapnia). The order of vasoconstriction and vasodilation was varied across sessions 2 and 3. The design of the protocol was simplified for the subjects from the older age group. Each subject from the older group attended for one visit. O2 and CO2 were administered to the subjects using a face mask and sequential re-breathing circuit. The order of gas provocation was varied among the subjects (i.e. hyperoxia or hypercapnia first). For both groups, measurements of vessel diameter, centerline blood velocity and derived blood flow were acquired at each condition (i.e. baseline, during stabilized vasoconstriction, vasodilation, and recovery) at two discrete measurement sites along the supero-temporal arteriole. Results: The results of the repeated measures ANOVA showed a significant difference between the narrow and wide measurement sites for the younger group for flow (p??? 0.0003) and a significant influence of inspired gas provocation on flow for both protocols (p<0.0001). In addition, the interaction of measurement site and inspired gas provocation was significant (p<0.0001). The magnitude of retinal vascular reactivity showed a significantly greater blood flow response for the wide measurement site (p<0.0001). O2 provocation resulted in vasoconstriction that was still present up to 10 minutes after cessation of the stimulus (order effect of O2; p???0.046). No such order effect was apparent for CO2 provocation (order effect of CO2; p=0.352). The group mean blood flow Coefficient of Repeatability (COR) for the narrow measurement site was 0.74 ??l/min (relative to group mean flow of 4.85 ??l/min ?? SD 1.31) and for the wide measurement site was 1.49 ??l/min (relative to group mean flow of 11.29 ??l/min ?? SD 3.55). There was no difference between the young and the older age groups in retinal vascular reactivity for both the narrow (two-tailed Student t-test, p=0.8692) and wide (two-tailed Student t-test, p=0.2795) measurement sites. Conclusion: This study demonstrated that the magnitude of retinal vascular reactivity was greater for arteriolar measurement sites with wider baseline vessel diameters. In addition, it demonstrated that hyperoxic provocation resulted in a persistent vasoconstriction up to 10 minutes after cessation of the stimulus. The study demonstrated that the repeatability of retinal blood flow measurements in absolute terms is lower for smaller diameter vessels. Finally, this study also suggests that age does not affect the relationship between retinal vascular reactivity and vessel diameter.

Chronic traumatic encephalopathy (CTE) is a progressive neurodegenerative disease that is characterized by a tauopathy in the form of aggregates of hyperphosphorylated tau throughout the CNS. Individuals suffering from CTE experience many different symptoms that result in dementia and severe cognitive decline along with a heightened occurrence of suicide. Repetitive mild traumatic brain injuries, such as concussions or subconcussive blows, are responsible for the development of CTE. Therefore, individuals who participate in contact sports, such as football and hockey, are at a high risk of developing this disease. Currently, there is no method for detecting CTE during life and the diagnosis can only be made at autopsy. At present nothing is known about the eye pathology of CTE. But if a distinctive profile of ocular abnormalities could be identified, it would raise the possibility that CTE could be diagnosed during life by an eye exam. For this research, ten eyes of individuals suffering from varying stages of CTE were collected, dissected, and observed under a microscope to find pathology associated with CTE. The antibodies of interest are pTDP-43, p62, αβ crystallin, and CP13. Pathology was found in the retina, mostly in the ganglion cell layer, throughout the different stages of CTE with the most severe pathology occurring in the most severe cases. These results can serve as a foundation for continued CTE research in the eye and ultimately result in potential ophthalmological diagnostic tests in individuals suffering from CTE.

by Sek Chung Fung. / Thesis (M.Phil.)--Chinese University of Hong Kong, 1980. / Bibliography: leaves 89-98.

No description available.

No description available.

Retinal imaging with a fundus camera or scanning laser ophthalmoscope (SLO) allows a fast, non-invasive view to the body’s microvasculature. Evidence suggests that features associated with retinal blood vessels (for example, narrowing of arteries and increased vascular tortuosity) measured near to the optic disc are early biomarkers of disease such as diabetes, hypertension (HT) and cardiovascular disease. Ultra-widefield (UWF) ophthalmic imaging allows unique views of peripheral locations as well as the posterior pole, potentially facilitating a more comprehensive study of the state or health of the microvasculature than is afforded by conventional retinal imaging. It is envisaged that this will reveal new candidate biomarkers derivable from the retina which could help identify the presence of disease or improve risk stratification for serious illness. In this thesis, 532 individuals were recruited from a trial evaluating the added benefit of using computed tomography (CT) imaging in the diagnosis of coronary heart disease (CHD) to measure atherosclerotic plaque in the arteries of the heart muscle tissue. The trial participants were deeply phenotyped which allowed access to additional information including: presence and severity of CHD, hypertensive status, presence of diabetes, age, gender, and smoking status (all risk factors commonly associated with CHD). After CT imaging patients were invited for undilated UWF ophthalmic imaging using an Optos P200C SLO. To accurately measure features of blood vessels indicative of microvascular health or disease in these images required the development of a novel semi-automatic computerised technique to segment and analyse the retinal vasculature. This involved implementation of a supervised vessel segmentation algorithm utilising multi-scale matched filters, a neural network classifier and hysteresis thresholding. A true positive rate (TPR) of 0.702 (and standard deviation of 0.059), false positive rate (FPR) of 0.011 (0.006) and accuracy (Acc) of 0.965 (0.006) was achieved by the algorithm. This compared to TPR of 0.674 (0.062), FPR of 0.017 (0.004), and Acc of 0.957 (0.006) for state-of-the-art fundus camera vessel segmentation applied to UWF SLO. After segmentation and prior to the measurements of retinal vessel parameters, the distortions introduced by the instrument mapping the 3D retinal surface onto a 2D image plane were also accounted for utilising an established technique. This is especially important for measuring in the periphery of UWF images and has not previously been reported for biomarker discovery. Measurements from UWF SLO were compared between those participants with CHD (where a reduction in arterial width was hypothesised based on existing research with fundus cameras into cardiovascular disease), and those without to investigate whether a difference between the two groups existed. After appropriate statistical correction for confounding variables (i.e. age, gender, and hypertensive status) the results did not show a statistically significant difference for presence of CHD or for risk stratification. However, the analysis techniques that were developed in this thesis do allow a rapid investigation of retinal vascular parameters in UWF SLO. This has application to a number of other diseases, such as HT, where a more pronounced change to the appearance of vessels is anticipated, and to different areas of the peripheral retina not previously measurable with standard imaging techniques and existing algorithms.

Leung Yiu Fai Ivan. / Thesis (M.Phil.)--Chinese University of Hong Kong, 1997. / Includes bibliographical references (leaves 41-46). / Acknowledgements --- p.ii / Table of contents --- p.iii / List of tables --- p.vi / List of figures --- p.vii / List of Abbreviations --- p.xii / Abstract --- p.xiii / Chapter I. --- INTRODUCTION --- p.1 / Chapter A. --- Current knowledge on carotenoids --- p.1 / Chapter 1. --- Chemistry --- p.1 / Chapter 2. --- Occurrence --- p.3 / Chapter 3. --- Metabolism --- p.6 / Chapter 4. --- Biological function --- p.8 / Chapter B. --- Statement of the problem --- p.10 / Chapter II. --- MATERIAL AND METHODS --- p.11 / Chapter A. --- Materials --- p.11 / Chapter 1. --- Animals --- p.11 / Chapter 2. --- Human samples --- p.11 / Chapter 3. --- Chemicals --- p.12 / Chapter 4. --- HPLC Apparatus --- p.12 / Chapter B. --- Methods --- p.14 / Chapter 1. --- Animal specimens --- p.14 / Chapter (a) --- Carotenoid supplement --- p.14 / Chapter (b) --- Tissue preparation --- p.14 / Chapter 2. --- Human specimens --- p.15 / Chapter 3. --- Extraction of lipid component --- p.15 / Chapter (a) --- Retina --- p.15 / Chapter (b) --- Serum or subretinal fluid --- p.16 / Chapter (c) --- Liver --- p.16 / Chapter 4. --- Analytical methods --- p.17 / Chapter (a) --- Isocratic elution --- p.17 / Chapter (b) --- Gradient elution --- p.17 / Chapter III. --- RESULTS --- p.18 / Chapter A. --- Selection of chromatographic method for carotenoid analysis --- p.18 / Chapter 1. --- Effect of dioxane concentration on the retention time of carotenoids --- p.18 / Chapter (a) --- Gradient elution --- p.18 / Chapter (b) --- Isocratic elution --- p.18 / Chapter 2. --- Chromatograms of carotenoids and retinoids of a selected method for routine analysis --- p.19 / Chapter (a) --- Carotenoids --- p.20 / Chapter (b) --- Retinoids --- p.21 / Chapter B. --- Application of the selected method to study carotenoid in human and rats --- p.22 / Chapter 1. --- Study in human serum and subretinal fluid --- p.22 / Chapter (a) --- Serum --- p.22 / Chapter (b) --- Subretinal fluid --- p.22 / Chapter 2. --- Study in rats tissues --- p.24 / Chapter (a) --- Liver --- p.24 / Chapter (b) --- Serum --- p.26 / Chapter (c) --- Retina --- p.28 / Chapter C. --- Influence of dietary carotenoids on retinol concentration in rats --- p.31 / Chapter 1 --- Serum --- p.31 / Chapter 2 --- Retina --- p.31 / Chapter IV. --- DISCUSSION --- p.32 / Chapter A. --- Chromatographic analysis of carotenoids and retinoids --- p.32 / Chapter B. --- Carotenoid study in human and rats --- p.33 / Chapter 1. --- Carotenoids in human tissues --- p.33 / Chapter (a) --- Serum --- p.33 / Chapter (b) --- Subretinal fluid --- p.34 / Chapter 2. --- Dietary supplement of carotenoids to rats --- p.35 / Chapter (a) --- Choice of animal and types of carotenoids --- p.35 / Chapter (b) --- Carotenoids uptake into rats tissues --- p.36 / Chapter (c) --- Effect of dietary carotenoids on retinol concentration in rat tissues --- p.38 / Chapter V. --- CONCLUSION --- p.39 / Chapter VI. --- REFERENCES --- p.41 / Chapter VII. --- TABLES --- p.47 / Chapter VIII. --- FIGURES --- p.56

LMO4 is a transcription cofactor expressed during retinal development and in amacrine neurons at birth. A previous study in zebrafish reported that morpholino RNA ablation of one of two related genes, LMO4b, increases the size of the eye in embryos. However, the significance of LMO4 in mammalian eye development and function remained unknown since LMO4 null mice die prior to birth. We observed the presence of a smaller eye and/or coloboma in ~40% of LMO4 null mouse embryos. To investigate the postnatal role of LMO4 in retinal development and function, LMO4 was conditionally ablated in retinal progenitor cells using the Pax6 alpha-enhancer Cre/LMO4flox mice. We found that these mice have fewer Bhlhb5-positive GABAergic amacrine and OFF-cone bipolar cells. The deficit appears to affect the postnatal wave of Bhlhb5+ neurons, suggesting a temporal requirement for LMO4 in retinal neuron development. In contrast, cholinergic and dopaminergic amacrine, rod bipolar and photoreceptor cell numbers were not affected. The selective reduction in these interneurons was accompanied by a functional deficit revealed by electroretinography, with reduced amplitude of b-waves, indicating deficits in the inner nuclear layer of the retina. Thus, LMO4 is necessary for normal GABAergic amacrine and OFF-cone bipolar cell development during retina development.

Inherited photoreceptor degenerations (IPDs) are the most common monogenic cause of blindness in humans. To discover genes that may influence the risk of death in IPDs, microarray studies were used, and ET-2 was identified as the most differentially expressed transcript. ET-2 mRNA was 32-fold (p< 0.004), 70-fold (p< 0.009) and 72-fold (p<0.0009) increased in the Rds+/- , Tg(RHO P347S) and Rd1-/- mouse models of IPD, respectively, and the ET-2 peptide was minimally three-fold upregulated in Rds+/- retinas. The increased ET-2 transcript was detected solely in the photoreceptors (PRs) of Rds+/- and Tg(RHO P347S) retinas, but not in wild-type (wt) retinas by in situ hybridization. To determine the biological role of ET-2 in IPDs, mouse IPD models were crossed to ET-2-null mice. At age 40 and 15 days, ET-2-/-; Tg(RHO P347S) and ET-2-/-;Rd1-/- retinas showed, respectively, a 41% (n=6;p<0.003) and 49% rescue of PR degeneration (n=5;p<0.007). Unexpectedly, however, the PRs in ET-2-/- Rd1-/-retinal explants were not rescued (n=6;p>0.05), suggesting that the rescue observed in vivo might be due to extraocular mechanisms. Additionally, the expression of ET-2 mRNA from an rAAV5-CBA-ET-2 vector in ET-2-/-; Rd1-/- retinas did not restore PR degeneration (n=6;p>0.05). A survey of the extraocular phenotypes of ET-2 null mice showed them to be hypoxic owing to aberrant lung development, with a loss of normal alveolarization of the lung. Erythropoietin (EPO) levels were 11-fold elevated in the serum of ET-2 null mice (n=7;p<0.05) and retinal vascular endothelial growth factor (VEGF) was increased 4-fold (n=4;p<0.05). To examine the role of hypoxia in PR degeneration and to exclude increased EPO levels as the sole factor accounting for the rescue of mutant PRs in ET-2-/-; Tg(RHO P347S) and ET-2-/-;Rd1-/- mice in vivo, the effect of hypoxia on PR death in Rd1-/- retinal explants was examined. Rd1-/- explants cultured in 6% O2 from PN10 to PN17 showed a 32% rescue of PR death (n=5;p<0.05). Although ET-2 may mediate PR death through a direct role in mutant PRs, the PR rescue observed in ET-2-/-; Tg(RHO P347S) and ET-2-/-;Rd1-/-retinas may also result from systemic hypoxia due to poor lung function in ET-2-/- animals.

Inherited photoreceptor degenerations (IPDs) are the most common monogenic cause of blindness in humans. To discover genes that may influence the risk of death in IPDs, microarray studies were used, and ET-2 was identified as the most differentially expressed transcript. ET-2 mRNA was 32-fold (p< 0.004), 70-fold (p< 0.009) and 72-fold (p<0.0009) increased in the Rds+/- , Tg(RHO P347S) and Rd1-/- mouse models of IPD, respectively, and the ET-2 peptide was minimally three-fold upregulated in Rds+/- retinas. The increased ET-2 transcript was detected solely in the photoreceptors (PRs) of Rds+/- and Tg(RHO P347S) retinas, but not in wild-type (wt) retinas by in situ hybridization. To determine the biological role of ET-2 in IPDs, mouse IPD models were crossed to ET-2-null mice. At age 40 and 15 days, ET-2-/-; Tg(RHO P347S) and ET-2-/-;Rd1-/- retinas showed, respectively, a 41% (n=6;p<0.003) and 49% rescue of PR degeneration (n=5;p<0.007). Unexpectedly, however, the PRs in ET-2-/- Rd1-/-retinal explants were not rescued (n=6;p>0.05), suggesting that the rescue observed in vivo might be due to extraocular mechanisms. Additionally, the expression of ET-2 mRNA from an rAAV5-CBA-ET-2 vector in ET-2-/-; Rd1-/- retinas did not restore PR degeneration (n=6;p>0.05). A survey of the extraocular phenotypes of ET-2 null mice showed them to be hypoxic owing to aberrant lung development, with a loss of normal alveolarization of the lung. Erythropoietin (EPO) levels were 11-fold elevated in the serum of ET-2 null mice (n=7;p<0.05) and retinal vascular endothelial growth factor (VEGF) was increased 4-fold (n=4;p<0.05). To examine the role of hypoxia in PR degeneration and to exclude increased EPO levels as the sole factor accounting for the rescue of mutant PRs in ET-2-/-; Tg(RHO P347S) and ET-2-/-;Rd1-/- mice in vivo, the effect of hypoxia on PR death in Rd1-/- retinal explants was examined. Rd1-/- explants cultured in 6% O2 from PN10 to PN17 showed a 32% rescue of PR death (n=5;p<0.05). Although ET-2 may mediate PR death through a direct role in mutant PRs, the PR rescue observed in ET-2-/-; Tg(RHO P347S) and ET-2-/-;Rd1-/-retinas may also result from systemic hypoxia due to poor lung function in ET-2-/- animals.