Inhibitory effect ofinsulin-like growth factor binding protein-related protein 1 on retinal angiogenesis invitro

Abstract

：Background Antagonistsagainst vascular endothelial growth factor (VECF) play key roles in treating andpreventing neovascular ophthalmopathy. As a novel anti-angiogenic factor, insulin-likegrowth factor binding protein-related protein 1 (IGFBP-rP1) might be an antagonist againstVEGF in eye. Objective This study was to explore the inhibitory effect of IGFBP-rP1, anovel anti-angiogenic factor, on VEGF-induced retinal angiogenesis in vitro. Methods Theretina-choroid endothelial cell line ( RF/6A ) was cultured in DMEM containing 10% fetalbovine serum. Culture cells were divided into control group(free-serum culture group),10mg/L VEGF culture group and different concentrations of IGFBP-rP1 (50,100,200 mg/L) +10mg/L VEGF group. The expression of IGFBP-rP1 in the cells was detected byimmunofluorescence assay. The proliferation and migration of RF/6A cells were evaluatedusing MTS colorimetric assay and the chemotactic motility assay, respectively. Flowcytometry was used to detect the apoptosis of RF/6A cells. Results The immunofluorescenceassay RF/6A cells showed the green fluorescence in cytoplasm and red fluorescence innuclei after cells were exposed to any concentration of IGFBP-rP1 ,but only redfluorescence was seen in nuclei in control cells. After stimulation of 10 mg/L VEGF,theproliferation value (A490) was elevated and the numbers of cell migration were increasedin comparison with control group (t = -15. 191, P = 0. 000; t = -21. 274, P = 0. 000 ) ,but the cellular apoptosis rate was lower than the control group (t - 10. 228, P = 0. 000) . After treated with various concentrations of IGFBP-rP +10% VEGF, the proliferation andmigration of RF/6A cells were significantly decreased in comparison with only 10% VEGFgroup (F = 534. 158,P = 0. 000;F = 2742. 323,P = 0.000,respectively) ,and the inhibitoryeffects were gradually enhanced with the increase of IGFBP-rP1 levels (P<0. 05). Theapoptosis rate of RF/6A cells in 50,100 and 200 mg/L + 10 mg/L VEGF groups increased by (1. 26±0. 04)% ,( 1. 50±0. 07)% and ( 1. 93±0. 27)% respectively,showing significantdifferences among different groups ( F = 274. 273, P = 0. 000). Conclusion IGFBP-rP1inhibits the proliferation and activity of retina and choroid endothelial cells induced byVEGF at a concentration-independent manner. It appears to be as a novel endogenousinhibitory factor in retinal angiogenesis. Key words: Insulin-like growth factor binding protein-related protein 1; Vascular endothelial growth factor; Retina; Vascular endothelial cell; Angiogenesis