Abstract
Purpose :
Age-related macular degeneration (AMD) is a chronic multifactorial disease characterized by an increase in oxidative stress, inflammation, maladaptive immune response, and dysregulated complement system pathways. Genetic variants within complement genes have been recognized in AMD, which influence the functionality of the complement system. Recent findings suggest that extracellular vesicles (EVs) play a role in AMD. Since EVs can travel through different biofluids, including plasma and serum, the present study aimed to analyze the cargo of circulating EVs in patients at different AMD stages.
Methods :
Total of 40 participants recruited from the University of Colorado AMD Registry were classified through multimodal imaging. Plasma samples from 10 control subjects, 10 intermediate AMD (iAMD) patients, 10 with Geographic Atrophy (GA), and 10 with Neovascular (NV) AMD were utilized for the EVs isolation by size exclusion chromatography. EVs characterization was performed considering their size, distribution, concentration release, and morphology by electron microscopy, and nanosight. EVs’ molecular cargo and purity were analyzed by western blot. We profiled EV content through proteomic analysis, emphasizing on the complement pathway. Finally, cross-validation of the results was performed using Western blotting and enzyme-linked immunosorbent assay (ELISA).
Results :
The isolated plasma-derived EVs exhibited a characteristic rounded membrane within a size range of 30–170 nm. Their physical characteristics and biochemical composition confirmed that they fulfilled the established criteria for EVs. Enriched pathways analysis of proteomic data significantly underscored the complement cascade. Circulating EVs from AMD patients carried complement proteins. The levels of complement proteins (C1q, C3b, and 5Cb-C9) and complement regulators (MBL2, KLKB1, CLU, and VTN) in plasma derived EVs were significantly different between AMD patients and healthy controls. Furthermore, different profiles were also observed when comparing iAMD stages and advanced AMD (GA and NV).
Conclusions :
Here we detected crucial proteins related to the complement system in plasma EVs derived from AMD patients enabling us to distinguish between AMD stages. The data suggest that we may be able to distinguish the different AMD phenotypes using EVs.
This abstract was presented at the 2024 ARVO Annual Meeting, held in Seattle, WA, May 5-9, 2024.