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S. Parameswaran, C. Spee, S. J. Ryan, R. Kannan, D. R. Hinton; Augmentation of Apical Secretion of VEGF by BMP-4 in Polarized ARPE-19 Cells. Invest. Ophthalmol. Vis. Sci. 2007;48(13):2962.
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Bone morphogenetic protein-4 (BMP-4), a member of the TGF-ß superfamily of proteins, plays an important role in embryonic development and is expressed in multiple tissues, including the retina. The aim of the present study was to investigate the effect of exogenous BMP-4 on VEGF secretion in ARPE-19 cells and characterize the polarity of secretion.
ARPE-19 cells were cultured for 1 month in DMEM/F12 medium containing 1% FBS on transwell filters to produce polarized RPE monolayers. Monolayer integrity was established by measuring transepithelial resistance (TER), and the presence of tight junctions (TJ) was assessed by ZO-1 and occludin and apical Na/K ATPase localization. Paracellular permeability was measured using [3H] mannitol. Human recombinant BMP-4 at concentrations of 10, 25, 50, 75 and 100 ng/ml was introduced to the apical and basolateral medium for 24 h. VEGF-A and C secretion to the apical and basolateral domains and cellular concentrations of VEGF were determined by ELISA.
Confocal microscopy confirmed the localization of TJ proteins ZO-1 and occludin and localization of Na/K ATPase at the apical surface of the plasma membrane. The TER values in ARPE-19 cells maintained for 1 month in 1% FBS-containing medium averaged 45.6 ± 5.2 Ω.cm2 and did not change significantly after BMP-4 treatment. No apparent significant difference in paracellular permeability of mannitol was observed between the two membrane domains. The cellular VEGF levels increased a maximal threefold with BMP-4 treatment for 24 h. The apical of secretion of VEGF-A and VEGF-C increased in a dose-dependent manner in polarized monolayers treated with BMP-4. The increase in VEGF-A and VEGF-C secretion was maximal with 100 ng/ml BMP-4 and was significant over controls (p<0.05).
Our results suggest that BMP-4 may play a role in the regulation of angiogenesis by stimulating release of VEGF-A and VEGF-C from RPE.
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