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R. Koyama, H. Cai, L.V. Del Priore; AGING OF BRUCH’S MEMBRANE DECREASES RETINAL PIGMENT EPITHELIUM (RPE) PHAGOCYTOSIS . Invest. Ophthalmol. Vis. Sci. 2004;45(13):3676.
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© ARVO (1962-2015); The Authors (2016-present)
Purpose: The earliest changes in age–related macular degeneration occur within Bruch’s membrane. We have previously shown that Bruch’s membrane aging affects the attachment and survival of the overlying RPE, and herein we determine the effects of Bruch’s membrane aging on RPE phagocytosis ability. Methods: Explants of human Bruch’s membrane were prepared from the periphery of donor human cadaver eyes (age range: 33– 43 for young group; 75 – 94 for older group) within 48 hours of death. 6 mm Bruch’s membrane explants were embedded with the basal lamina up in 96–well culture plate. 50,000 immortalized ARPE–19 harvested from confluent culture were seeded onto 96 well culture plates containing Bruch’s membrane explants and bare control wells for 72 hours; 1 ul of fluorescent latex beads (3.6x105 beads/ul) were added to each well for another 24 hours prior to trypsinizing the RPE. Ingested fluorescent beads were counted using a FACS Flow Cytometer. Results: The percentage of cells ingesting beads was consistently higher on bare plastic versus Bruch’s membrane (10.11 + 3.51 % vs. 4.10 + 1.88 %, respectively; p = 0.00004). When we considered only the population of cells that had ingested beads, the fluorescent intensity per cell, which is a measure of the number of beads ingested per cell, was similar between the two groups (p>0.05). The percent of cells ingesting beads on young Bruch’s membrane versus older Bruch’s membrane were similar (4.04 + 1.57 % vs. 4.14 + 2.19 %, respectively; p > 0.05). In contrast, the fluorescent intensity within the cells cultured on younger Bruch’s membrane was consistently higher than the fluorescent intensity on older Bruch’s membrane (312.80 + 36.82 % vs. 246.09 + 62.83 % respectively; p = 0.02). Conclusions: This study suggests that Bruch’s membrane inhibits RPE phagocytosis compared to bare tissue culture plastic, and aging of Bruch’s membrane decreases RPE phagocytosis further than young Bruch’s membrane. To our knowledge, this is the first demonstration that aging of Bruch’s membrane can modulate RPE phagocytosis ability. Further study is required to determine the implications of this age–dependent decrease in RPE phagocytosis in the pathogenesis of AMD.
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