The effects of recombinant human platelet derived growth factor-BB on biological behaviours of human retinal vascular endothelial cells

Abstract

Objective This study was to investigate the role of recombinant human platelet derived growth factor-BB(rhPDGF-BB) in the proliferation and migration of human retinal vascular endothelial cells (hRVECs). Methods hRVECs were cultured in DMEM with 10% fetal bovine serum.The rhPDGF-BB at the concentrations of 10, 50 and 200 ng/ml were added into the medium of exponential phase-growth cells for 24 and 48 hours, respectively, and no rhPDGF-BB was added in the normal control group.The proliferation of the cells (absorbancy) was assayed by cell counting kit 8(CCK8) method.Cell scratch test was employed to evaluate the relative migration area of cells (migrated acellular area/initial acellular area). The relative expression of rhPDGF-BB recepter (rhPDGF-BBR) mRNA in the cells was detected by reverse transcription PCR.The relative expression of VEGF mRNA and integrin mRNA in the cells was detected using real-time fluorescence quantitative PCR. Results hRVECs grew well and a expressing band according with rhPDGF-BBR prime was displayed.The absorbancy values of the cells were 1.01±0.05, 1.09±0.04, 1.10±0.02 and 1.13±0.05 in the normal control group and 10, 50, 200 ng/ml rhPDGF-BB groups at 24 hours after culture, and those in the 10, 50 and 200 ng/ml rhPDGF-BB groups were significantly increased in comparison with the normal control group (t=2.504, 3.430, 3.483, all at P<0.05). The relative migrated areas of the cells in the normal control group and 10, 50, 200 ng/ml rhPDGF-BB groups were 0.42±0.10, 0.38±0.09, 0.55±0.06 and 0.61±0.05 at 24 hours after culture, and those at 48 hours were 0.75±0.06, 0.81±0.02, 0.87±0.02 and 0.98±0.02, showing significant differences among the groups (Fgroup=16.283, P=0.000; Ftime=209.129, P=0.000), and the relative migrated areas was depended upon the rhPDGF-BB dose and time.The relative expressions of integrin mRNA were 1.06±0.02, 1.30±0.10, 1.20±0.16 and 1.27±0.08, and those of VEGF mRNA were 0.97±0.05, 1.06±0.16, 1.58±0.18 and 1.66±0.21 in the normal control group and 10, 50 ng/ml, 200 ng/ml rhPDGF-BB groups, respectively, and increased expressions of integrin mRNA and VEGF mRNA were found in the 50 and 200 ng/ml rhPDGF-BB groups compared with the normal control group (integrin mRNA: t=3.900, 4.014, both at P<0.05; VEGF mRNA: t=6.940, 7.210, both at P< 0.05). Conclusions rhPDGF-BB/rhPDGF-BBR signal promotes the proliferation and migration of hRVECs probably by up-regulating the expressions of integrin and VEGF. Key words: Endothelium, vascular/cytology; Neovascularization, pathologic/metabolism; Platelet-derived growth factor; Receptors, platelet-derived growth factor; Recombinant proteins; Humans; Cells, cultured; Integrin; Vascular endothelial growth factor