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Shawn A. Morales, David G. Telander, Deanna Leon, Madhuri Wadehra, Jonathan Braun, Lynn K. Gordon; Epithelial Membrane Protein 2 (EMP2) Control Of VEGF Expression In ARPE-19 Cells Is Functionally Significant. Invest. Ophthalmol. Vis. Sci. 2011;52(14):4857.
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Age-related macular degeneration (AMD) is the primary cause of blindness in people 50 and older in developed countries. VEGF production by retinal pigment epithelial cells has been shown to be a key molecule regulating aberrant angiogenesis in the retina. Epithelial membrane protein 2 (EMP2), a member of the tetraspan protein family, is highly expressed in the RPE and has been shown to regulate FAK activation. FAK signal transduction is implicated in VEGF production. The purpose of this study is to determine whether EMP2 regulates modulation of VEGF expression in RPE and whether those changes are functionally significant.
ARPE-19 cells were engineered to overexpress EMP2 (ARPE-19/EMP2). Both ARPE-19 and ARPE-19/EMP2 cells were evaluated for VEGF expression. In some experiments, the cells were pretreated with 20µg/ml of anti-EMP2 or control diabody for 2 hours. The small molecule inhibitor PP2 was used to inhibit FAK activation. VEGF levels were measured by Western Blot and ELISA. HUVEC migration was assayed in a transwell system.
VEGF expression increased by 150% in EMP2 overexpressing cells as compared to ARPE-19 cells (P=.003). Exposure to anti-EMP2 diabody resulted in a 75% reduction in EMP2 protein levels. Concordant decreases in basal levels of activated FAK were also observed. Anti-EMP2 diabody treatment reduced VEGF expression by 50% in ARPE-19 (P=.04) and by 40% in the ARPE-19/EMP2 cells(P=.03). VEGF secretion was reduced by 64% in cells treated with PP2. Increased expression of EMP2 resulted in a 3-fold increase in HUVEC migration.
This study establishes a novel connection between levels of EMP2 and VEGF and may reflect either a direct effect through the tetraspan web or an indirect change through FAK activation. This connection is functionally significant. In addition to the direct use of anti-VEGF antibodies, EMP2 may be an additional therapeutic target for the treatment of neovascular AMD.
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