Insulin-Like Growth Factor Binding Protein-Related Protein 1 Mediates VEGF-Induced Proliferation, Migration and Tube Formation of Retinal Endothelial Cells

Abstract

Purpose: The potential role of insulin-like growth factor binding protein-related protein 1 (IGFBP-rP1) as an endogenous angiogenesis inhibitor in the prevention of vascular endothelial growth factor (VEGF)-induced retinal angiogenesis has not been explored.

Methods: Expression of IGFBP-rP1 in rhesus macaque choroid-retinal endothelial cell line (RF/6A) cells was assessed by reverse transcription polymerase chain reaction analysis and Western blotting. RF/6A cells were treated with VEGF (10 ng/ml) alone or in the presence of IGFBP-rP1 at concentrations ranging from 50 ng/ml to 200 ng/ml. The proliferation, migration and capillary-like tube formation of RF/6A cells were evaluated using 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium colorimetric assay, the chemotactic motility assay and the Matrigel tube formation assay, respectively. Flow cytometry was used to detect the apoptosis of RF/6A cells, and oncogenic B-Raf expression was assessed to elucidate the pathway for IGFBP-rP1-mediated induction of apoptosis in the presence of VEGF.

Results: RF/6A cells expressed both IGFBP-rP1 transcripts and IGFBP-rP1 protein. VEGF markedly stimulated proliferation, migration and capillary-like tube formation of RF/6A cells (P<0.05), whereas those VEGF-induced parameters were significantly inhibited by IGFBP-rP1 at concentrations ranging from 50 ng/ml to 200 ng/ml in a dose-dependent manner (P < 0.05). Following addition of IGFBP-rP1, expression of B-Raf was significantly decreased dose-dependently, and apoptosis occurred as evidenced by flow cytometry (P < 0.05).

Conclusions: IGFBP-rP1 can inhibit the stimulatory effect of VEGF on retinal angiogenesis in vitro by inhibiting expression of B-Raf to induce apoptosis. It is a novel endogenous anti-angiogenic factor with potential therapeutic action in retinal neovascularization dependent disorders.

KEYWORDS::

• Angiogenesis
• B-Raf
• Insulin-like growth factor binding protein-related protein 1 (insulin-like growth factor binding protein-7)
• Retina
• Vascular endothelial growth factor

ACKNOWLEDGMENTS

This work was supported by grants from the State Key Program of National Natural Science Foundation of China (No. 30930097), the National Natural Science Foundation for Young Scholars of China (No. 30801269), the Doctoral Innovation Fund of Shanghai Jiao Tong University School of Medicine (No. BXJ0935), and the Program of Shanghai Key Laboratory of Ocular Fundus Diseases (No. 09-10).

Declaration of interest: The authors report no conflicts of interest. The authors alone are responsible for the content and writing of the paper.