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S. Sonoda, P. G. Sreekumar, C. Spee, S. J. Ryan, R. Kannan, D. R. Hinton; Selective Increase in Basolateral Secretion of VEGF Upon BMP-4 Treatment of Highly Polarized Human RPE Cells. Invest. Ophthalmol. Vis. Sci. 2008;49(13):855.
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Bone morphogenetic protein-4 (BMP-4) is a member of the TGF-beta superfamily and plays an important role in development, morphogenesis, cell proliferation and apoptosis. It is expressed in a number of tissues including the retina. We investigated the effect of exogenous BMP-4 on secretion of two key angiogenic growth factors, VEGF and PEDF, by the apical and basolateral domains of highly polarized human RPE.
Highly differentiated RPE cells were grown on Transwell filters (12 mm, 0.4µm pore size). RPE cells were cultured in a differentiation media modified from that previously described by Hu and Bok (Mol Vis. 2001; 7:14-9) for 1 month. The integrity and resistance of the RPE monolayer was evaluated by measuring transepithelial resistance (TER) and staining for tight junction (TJ) proteins. Human recombinant BMP-4 (10, 25, 50, 75 and 100 ng/ml) was introduced to the apical and basolateral transwells in serum-free medium. Cellular expression and secretion of VEGF-A and PEDF to the apical and basolateral domains were determined by ELISA after 24 h.
The integrity of RPE layer was confirmed by expression of TJ proteins ZO-1 and occludin and apical localization of Na/K-ATPase. The TER averaged 503.2 ± 128.4 ohms.cm2 and did not change after BMP-4 treatment. A dose-dependent increase in basolateral secretion of VEGF was observed after BMP-4 from 830.1± 232.7 pg/ml in untreated control to a maximal level of 3034.7± 78.6 with 100 ng/ml while the apical secretion showed no apparent change. PEDF secretion from both apical and basolateral domains was unaffected by BMP-4 treatment. The VEGF to PEDF ratios were 1.1, 1.7, 2.2, 2.4 and 3.0 with 10, 25, 50 and 75 and 100 ng/ml, respectively (Control ratio = 1). The intracellular VEGF and PEDF levels were not significantly altered by BMP-4 treatment.
Our data suggest that BMP-4 may play a significant role in the regulation of angiogenesis in RPE by stimulating VEGF secretion to the choroidal side and by augmenting VEGF/PEDF ratios.
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