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A. M. Chan, S. A. Morales, A. Nagy, J. Braun, L. K. Gordon; High Expression of Epithelial Membrane Protein 2 (EMP2) Decreases Phagocytosis by ARPE-19. Invest. Ophthalmol. Vis. Sci. 2010;51(13):483.
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© ARVO (1962-2015); The Authors (2016-present)
Circadian cycle-dependent phagocytosis of photoreceptor outer segments (POS) by retinal pigment epithelial (RPE) cells is essential for the mammalian visual system. Integrin αvβ5 and activated focal adhesion kinase (FAK) are crucial for POS phagocytosis. EMP2 regulates formation of α/β integrin heterodimers and activation of integrin-FAK signaling complexes. The goal of this study was to determine whether changes in EMP2 expression levels influence POS phagocytosis.
Wild type, high EMP2-expressing (ARPE-19/EMP2) and EMP2-knock down (ARPE-19/EMP2siRNA) cells were assayed for phagocytosis using labeled bovine POS. Bound and internalized POS were quantitated by immunfluorescence image capture (Ariol SL-50). Levels of αvβ5 integrin were detected by flow cytometry, immunofluorescence and immunoprecipitation followed by Western Blot analysis.
In comparison to the ARPE-19 cells, ARPE-19/EMP2 cells exhibited decreased POS phagocytosis (P<0.01). Concordantly, ARPE-19/EMP2siRNA cells showed increased POS phagocytosis (P<0.05). Analysis of the percent distribution of bound and internalized POS identified a significant reduction in binding of POS to the surface of high EMP2-expressing cells (P<0.01), but enhanced internalization (P<0.05) consistent with the enhanced FAK activation by EMP2 overexpression. β5 integrin and αvβ5 integrin complex expression is significantly decreased in the high EMP2 expressing cells (P<0.05).
Elevated EMP2 level significantly reduces the overall POS phagocytosis in cultured ARPE-19 cells by down-regulating αvβ5 integrin. The increased POS engulfment rate resulting from the enhanced activation of the downstream signal pFAK cannot compensate for this diminished binding capacity. Changes in EMP2 expression levels in RPE cells in vivo could potentially lead to alterations in phagocytosis of POS.
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