The extracellular vesicle marker, Alix, co-localizes with Fibulin-3 puncta located in human drusen deposits

Abstract

Purpose : Purpose: Fibulin-3 (Fib3) is a secreted glycoprotein that is expressed in the retina and has been associated with drusen formation in age-related macular degeneration (AMD). The purpose of this study was to assess whether Fib3 is associated with extracellular vesicles (EVs) in drusen from non-diseased and AMD human donors.

Methods : Methods: De-identified sections of human eyes were received from the National Disease Research Institute (NDRI, Philadelphia). Donor eyes were either non-diseased (no known ocular pathology) or had been diagnosed with AMD. Retinal cryostat sections were labeled with primary antibodies targeted to Fib3, Apolipoprotein E (ApoE; a drusen marker), and ALG-2 interacting protein X (Alix, an EV marker) for confocal imaging (Leica TCS SP8).

Results : Results: Fib3 puncta was detected on the apical region of the RPE layer and within large AMD drusen. Alix-positive puncta was also detected in AMD drusen, where a number were Fib3-positive and the remaining were Fib3-negative. Similarly, there were Fib3-positive puncta that were Alix-negative. Fib3 and Alix also showed a degree of colocalization in the photoreceptor outer segments of the neural retina.

Conclusions : Conclusions: Our data suggests that the Alix-positive puncta are EV-rich populations which are accumulated, together with Fib3, within the drusen matrix during AMD. The EV populations possess a heterogeneous cargo content i.e. some carry the Fib3 protein and some do not. This shows a potential pathological role for Fib3 in AMD drusen formation which may utilize RPE-derived EVs for delivery.

This is a 2020 ARVO Annual Meeting abstract.