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Kieran S. Boochoon, Josh T. Davis, Joseph C. Manarang, Alison M. McDermott, William J. Foster; The Influence of Substrate Elastic Modulus on Retinal Pigment Epithelial Cell Phagocytosis. Invest. Ophthalmol. Vis. Sci. 2012;53(14):6567.
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The retinal pigment epithelial (RPE) cells lie on Bruch's membrane, directly under the retina, and phagocytose shed photoreceptor outer segments. Bruch's membrane is known to increase in stiffness by an order of magnitude (from 1,000 Pa to 10,000 Pa) with age. To better understand whether phagocytosis is influenced by substrate stiffness, we investigated the influence of substrate elastic modulus on the RPE cell line ARPE-19.
ARPE-19 cells were plated on 1mg/ml laminin-coated polyacrylamide substrates of varying elastic modulus (500 Pa, 1000 Pa, and 5000 Pa) and on a laminin-coated glass control. After 14 days in culture, 4,733 FITC-labeled latex fluorescent beads (diameter 1.967µm) suspended in PBS were placed in each well. After an incubation time of 4 hours, flow cytometry was performed to determine the distribution in the number of beads phagocytosed per cell.
Per 10,000 ARPE-19 cells grown on a 500Pa, 1000Pa, and 5000Pa elastic modulus, each set of ARPE-19 cells phagocytosed 2,631±51, 3,395±58, and 3,618±60 latex fluorescent beads respectively. The number of latex fluorescent beads phagocytosed by ARPE-19 increased continuously as a function of increasing substrate elastic modulus (p=0.0327; ANOVA).
Our results suggest that RPE cells display increased phagocytosis when grown on firmer substrates and thus RPE cells in older eyes, in which Bruch’s membrane is stiffer, may demonstrate increased phagocytosis. Drusen formation is believed to be triggered by photoreceptor byproducts that are incompletely metabolized by the retinal pigment epithelium. Impaired phagocytosis in RPE cells may contribute to impaired metabolism of photoreceptor outer segments and to the formation of drusen.
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