b isoforms induce proliferation of endothelial cells in culture through VEGFRs activation, although to a lesser extent than VEGF
. A: MTT assays. Commercial recombinant VEGF165 (R&D systems), VEGF121b and VEGF165b isoforms produced in P. pastoris (pp), and VEGF165b produced in mammalian cells (hs) were added to HUVECs alone or in combination. VEGF165 induces proliferation (p < 0.01) of HUVECs by 63% compared to control untreated cells. Co-administration of VEGF121b (pp), VEGF165b (pp) or VEGF165b (hs) with VEGF165 does not abrogate this effect. Addition of VEGF121/165b isoforms alone induces proliferation of HUVECs by ~40% over untreated cells. Co-treatment with VEGF165b (hs) and the VEGFR inhibitor GW654652 (1 μM) blocks the effect on proliferation. B: Analysis of DNA synthesis by incorporation of the modified nucleotide EdU. Administration of bFGF and VEGF165 increases DNA incorporation into HUVECs by 3-fold compared to untreated controls. Exposure to VEGF165b(pp) and VEGF121b(pp) also increases significantly DNA incorporation (by almost 2-fold) as compared to controls. *: p < 0.05; **: p < 0.01; ***: p < 0.001.