RNA-seq of human umbilical vein endothelial cells (HUVECs) stimulated with recombinant Vascular Endothelial Growth Factor A (VEGF-A) against untreated control

In order to identify new genes potentially involved in angiogenesis process, we performed a deep-sequencing transcriptome analysis of HUVECs stimulated with recombinant VEGF-A, introducing a difference with respect to the several previous studies. Primary ECs are normally cultured in vitro using an endothelial growth medium (EGM) constituted by an endothelial basal medium (EBM) supplemented with 2% fetal bovine serum (FBS) and a mix of growth factors and reagents, typically represented by basic Fibroblast Growth Factor (bFGF), Insulin-like Growth Factor 1 (IGF1), Epidermal Growth Factor (EGF), heparin, hydrocortisone, ascorbic acid and the same VEGF-A. In previous studies, HUVECs or other primary ECs were starved in EBM with low FBS concentration before the stimulation with VEGF-A. Instead of starvation, we grown HUVECs in complete EGM-2 medium except for VEGF-A to maintain the best condition for in vitro survival and propagation of primary ECs. After 24 hours of VEGF-A deprivation, VEGF-A was re-added to the medium and the stimulation was performed for six hours.