HUVEC cell wound scratch assay

HUVEC cells were seeded in six-well plate pre-coated with 0.1% gelatin in complete medium, after cells reached 70-80% confluence as a monolayer, cells were treated with 10 μg/ml mitomycin for 2 h to block cell proliferation and then the monolayer was scratched with a new 10 μl pipette tip across the center of the well. After scratching, the detached cells were removed by gently washing the well with culture medium twice. ECGM containing 0.5% FBS was added with or without 20 ng/mL VEGF and different concentrations of eupafolin. 12 h later, randomly chosen fields were photographed and the migrated cells were quantified by manual counting. Inhibition percentage was calculated by taking VEGF-induced wells as 100%.