Using an automated ocular vascular analysis software tool to evaluate the role of the inner blood retinal barrier in AMD pathogenesis and examining its application in neurological disorders

Abstract

Despite being the most common cause of central retinal blindness globally, there is no treatment for end stage dry age related macular degeneration (AMD). The prevailing school of thought with regard to AMD pathogenesis is that it is a disease of the outer retina. Macular translocation surgeries demonstrate disease recurrence in the geographic atrophied retina, implicating the inner retina in AMD pathogenesis. Hudson et al investigated the properties of the inner retinal vasculature integrity and demonstrated a circadian entrainment in animal and human studies (Hudson et al., 2019). The work from this group led to the establishment of the Irish Retinal Circadian Project (IRCP) which is a longitudinal study now in its second year, seeking to ascertain normal retinal interstitial kinesis with respect to circadian rhythms. The work has led to the generation of a dataset of young healthy control adults and age matched controls which provides researchers with a reliable model of normal inner retinal vascular homeostasis. Hudson et al previously demonstrated that the tight junction protein claudin-5 which is richly expressed at the inner blood retinal barrier (iBRB), is under the control of circadian clock component BMAL-1 (Hudson et al., 2019). Animal and human studies demonstrated a circadian entrainment to inner retinal vasculature permeability though fundus fluorescein angiography (FFA) studies (Hudson et al., 2019). It is widely accepted that many neurodegenerative diseases are associated with circadian rhythm disarray. Subsequently, we are interested in assessing the integrity of this circadian entrainment in a neurodegenerative disorder of the eye- AMD. Over the past two years, the group has turned to assess the effect of aging and neurodegeneration on iBRB behaviour. This project sought to build on the previous work of Hudson et al and expand the AMD dataset. We recruited subjects with all stages of dry AMD to undergo FFA scans in the morning and evening. We also recruited young healthy controls and age matched controls to undergo these same investigations to add to our dataset. Loss of integrity of the iBRB gives rise to a diseased state. Therefore, it follows that the quantification of the extent of this permeability may be of academic and clinical use. To fully ascertain the role that the iBRB plays in AMD, the degree to which the integrity of the iBRB is lost must be established. This logic led to the development of "Fluorescent Ocular Vasculature Analysis Software" ("FOVAS"), an automated quantification analysis software tool for FFA studies. FOVAS was employed in the analysis of the FFAs acquired in the morning and evening from AMD patients and controls. This project outlines FOVAS development but also demonstrates the role that FOVAS has in research methodologies. We demonstrate the applications of FOVAS in AMD research under the theory of iBRB disruption as an initiating event in AMD pathogenesis. We sought to characterise the effect of circadian cycling, aging and disease on retinal vasculature behaviour. However, the tight junction protein claudin-5 which is richly expressed at the iBRB is also expressed at the level of the blood brain barrier(BBB). Therefore, it follows that diseases affecting the BBB may also manifest themselves in the more readily accessible inner retina. This provides an excellent opportunity for the development of a diagnostic and prognostic tool for clinicians by way of FFA scans, even outside of ophthalmology. The IRCP control dataset provides a valuable insight into normal inner retinal function which can be useful in the comparison of diseased states. We therefore also sought to investigate the application of FOVAS in a number of neurological conditions that we hypothesized might have iBRB disruption. We report increased permeability of the iBRB in a single case of an individual with a condition in involving a dominant acting mutation in the gene CSF1R and in a small case series of patients with Visual Snow syndrome following FOVAS analysis. We conclude that with the development of FOVAS and a robust dataset demonstrating "normal" inner retinal vascular integrity, we can potentially expedite the study of blood tissue barrier disruption in neurodegenerative diseases. We hope that FOVAS will facilitate a new understanding of neurovascular physiology and help in the identification of a therapeutic target for neurodegenerative diseases.