Abstract
Purpose: :
VEGF-A plays an important role in angiogenesis, binding FLT1 (VEGFR1) or KDR(VEGFR-2). Previous studies suggest that KDR activation precedes angiogenesis, while FLT1 activation inhibits angiogenesis. In laser induced choroidal neovascularization (CNV), as a model of CNV to mimic age-related macular disease (AMD), SPARC (secreted protein, acidic and rich in cysteine) regulates whether VEGF-A binds to FLT or KDR. In this study, we attempted to control SPARC activity using anti-SPARC peptide to reduce laser induced CNV in mice.
Methods: :
Anti-SPARC peptide (amino acid sequence, KFMDVYNR) was created by phage display screening. Binding constants between anti-SPARC peptide and SPARC were measured by Biacore core facility at the University of Utah. Mouse eyes were injected with anti-SPARC peptide (20ng or 2ug in PBS) or PBS. One day after injection laser induced CNV was performed on all injected eyes. Ten days after laser induced CNV, eyes were enucleated, fixed in 4% PFA, and areas of CNV were stained with isolectin GS-IB4, and imaged using a confocal microscope. The volume of the areas of laser induced CNV were determined.
Results: :
The binding constant between anti-SPARC and SPARC was 5.0 x 102 M. Although the binding constant was low, intravitreal injection of 20ng and 2ug anti-SPARC peptide one day before laser injury showed decrease of laser CNV volume compared to PBS injection. Mean ±SEM CNV volume of eye in groups injected with PBS was 2.7 x 105(±4.2 x 104) um3, and eye injected with 20ng and 2ug anti-SPARC peptide was 1.4 x 105(±5.3 x 104) um3 (p=0.037) and 1.2 x 105(±5.1 x 104) um3 (p=0.026).
Conclusions: :
Inhibition of SPARC by anti-SPARC peptide reduces laser CNV volume in mice. These results indicate that SPARC has a significant role in CNV development, and could be an anti-angiogenic target for the treatment of AMD.
Keywords: choroid: neovascularization • vascular endothelial growth factor