The in vitro permeability assay was performed as we previously described [34 (link)]. Briefly, hBMECs transfected with miR-24 mimic or miR scramble were grown on 0.4-mm fibronectin-coated (R&D Systems) Transwell filters (Corning Inc., Corning, NY, USA). After 48 hours, the medium in the upper well was replaced by FITC-dextran 70 kD (0.5 mg/ml in PB).
Cells were stimulated in the lower well with PBS alone or PBS containing 50 ng/ml VEGF-A165 (R&D Systems, Inc., Minneapolis, MN). The entity of endothelial permeabilization was determined measuring at 520 nm the fluorescence of Dextran that passed in the bottom chamber through the cell monolayer.
Cells were stimulated in the lower well with PBS alone or PBS containing 50 ng/ml VEGF-A165 (R&D Systems, Inc., Minneapolis, MN). The entity of endothelial permeabilization was determined measuring at 520 nm the fluorescence of Dextran that passed in the bottom chamber through the cell monolayer.
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