A mouse model to study Aβ-driven pathology in the ageing retina

Lynn, Savannah, Goverdhan, Srini, Munday, Rosie, Scott, Jennifer, Freeman, Thomas, Johnston, David, Lotery, Andrew and Ratnayaka, J. Arjuna (2016) A mouse model to study Aβ-driven pathology in the ageing retina. Investigative Ophthalmology & Visual Science, 57 (12), 6534.

Abstract

Purpose : The Alzheimer’s-linked Amyloid beta (Aβ) peptide is reported to be deposited in aged retinas. Aβ has been implicated in key stages of Age-related Macular Degeneration (AMD), yet its role remains poorly understood. Here we employ a mouse model to study the in-vivo effects of Aβ to delineate its mechanisms of action and to understand how Aβ triggers/drives retinal pathology with age. Methods : Aβ was characterised by negative stain TEM and immunogold labelling. C57BL/6 mice were subretinally injected with 3µL of oligomeric Aβ1-42 (625nM, n=5) or vehicle control (n=3). At 8 days post-injection eyes were enucleated, OCT-embedded and cryosectioned at 16µM intervals for histological analysis. H&E staining and confocal immunofluorescence analysed retinal morphology in response to Aβ exposure and reported Aβ localisation. Exclusion criteria included a 200µM radius from the injection site to omit areas of mechanical trauma. Morphometric analysis was performed blind using OlyVIA and ImageJ. Data is expressed as means ± SEM with a statistical significance of *P ≤ 0.05. Results : TEM and Dot Blot assay enabled us to identify a window in which Aβ is reported to be most toxic. Fundus images showed large areas of pathology in Aβ exposed mice (9582 ± 4831) compared to controls (48.33 ± 8.97) which were indistinguishable from non-injected littermates, p=0.12. Serial line scans of Aβ injected mice revealed a 2-fold increase in RPE hypopigmentation associated with photoreceptor outer segment (POS) loss, RPE disorganisation and RPE hypertrophy compared to controls. Similarly, confocal data showed POS and inner segment disorganisation compared to well-preserved retinal architecture in control retinas. Aβ was detected in multiple retinal locations including POS, the RPE/choroid interface and the outer and inner plexiform layers. Conclusions : Our findings demonstrate that subretinal Aβ injections faithfully recapitulate key features of early AMD including dysfunctional RPE and damaged photoreceptors. Critically, we found no obvious indication of apoptosis or disruption of the blood-retinal barrier indicating gradual cellular impairment over time. Previously published literature showing static images of Abeta in human post-mortem eyes tantalisingly correlates the presence of Aβ with high drusen-loads and AMD. Our model therefore represents a powerful tool to investigate the dynamic nature of Aβ-mediated pathology in living retinas.

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Contributors

Author: Savannah Lynn

Author: David Johnston

Author: Andrew Lotery

Author: J. Arjuna Ratnayaka

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