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K. Sun, H. Cai, L.V. Del Priore; Bruch’s Membrane Aging Decreases Phagocytosis of Outer Segments by Retinal Pigment Epithelium . Invest. Ophthalmol. Vis. Sci. 2005;46(13):4165.
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Purpose: We have previously shown that Bruch’s membrane aging affects the attachment and survival of the overlying retinal pigment epithelium (RPE) and the ability of human RPE to ingest fluorescently labeled beads. Here we determine the specific effects of Bruch’s membrane aging on RPE phagocytosis of rod outer segments (ROS). Methods: Explants of human Bruch’s membrane were prepared from the periphery of donor human cadaver eyes (ages: 16 –74) within 48 hours of death. 6 mm Bruch’s membrane explants were embedded with the basal lamina up in 96–well culture plate. ARPE19 were seeded onto the explant surface (50,000 cells/ well) and cultured for one week. ROS isolated from adult bovine eyes were purified by sucrose gradient centrifugation, labeled with 10ug/ml fluorescein isothiocyanate and added to RPE on Bruch’s membrane (1.0 x 107 ROS/ml for 24 hr). RPE were examined by fluorescence microscopy and phagocytic activity was quantified by flow cytometry of harvested cells. Results: There was a significant difference in the ingestion of fluorescently–labeled ROS in RPE cultured on younger versus older Bruch’s membrane (873 + 9.3 vs. 608 + 25.4 arbitrary intensity units, respectively; p <0.01). The percentage of cells ingesting labeled ROS was similar in RPE on younger versus older BM (96% versus 88%, respectively). Conclusions: Aging of human Bruch’s membrane decreases RPE phagocytosis of ROS. To our knowledge, this is the first demonstration that aging of Bruch’s membrane can modulate ROS–specific phagocytosis. This observation is consistent with the hypothesis that a biologically–significant age–dependent decrease in RPE phagocytosis may play a role in the pathogenesis of aging disorders, including macular degeneration.
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