IGFBP-3 Acts as a Hematopoietic Stem Cell Homing Factor in an Adult Mouse Ocular Neovascularization Model

Abstract

Purpose: : IGFBP-3 has been shown to have direct pro-angiogenic effects on bone marrow derived hematopoietic stem cells and endothelial progenitor cells (EPC’s). IGFBP-3 stimulates HSC differentiation into endothelial cells and promotes their migration and tube formation in vitro. IGFBP-3 protects the retinal vasculature from oxygen induced damage by promoting vascular stability during hyperoxia exposure and vascular re-growth following return to room air in the oxygen induced retinopathy (OIR) model. In this study, we asked whether IGFBP-3 can also act as a homing factor for circulating HSC’s/EPC’s to establish residence in the retina. To test this we used an adult rodent ocular neovascularization model of laser induced retinal vessel occlusion.

Methods: : GFP⁺ chimeric mice were generated by isolating Sca-1⁺/c-Kit⁺ cells from GFP⁺ male mice via FACS sorting. Approximately 5,000 Sca-1⁺/c-Kit⁺ cells were injected into the retro-orbital sinus of 12 female mice. Stable engraftment was confirmed by flow cytometry 2 months following cell transplantation. The GFP⁺ stable chimeric mice were randomized into three treatment groups, laser only (n=4), IGFBP-3 injection only (n=4), and laser immediately followed by intravitreal injection of IGFBP-3 plasmid (n=4) (2ul of IGFBP-3 plasmid prepared in liposomes). The mice were sacrificed 3 weeks later. Flatmounts of the neural retina were prepared for retinal vessel visualization by immunohistochemisty and confocal microscopy.

Results: : Lasered mice treated with IGFBP-3 displayed large numbers of GFP⁺ cells incorporated into retinal vessels compared to fellow control eyes. Similarly mice treated with IGFBP-3 alone showed abundant numbers of GFP⁺ cells integrated into the retinal vessels even in the absence of laser induced retinal injury.

Conclusions: : IGFBP-3 can act as a HSC/EPC homing factor in an adult mouse neovascularization model. HSC’s can incorporate into retinal vessels and participate in repairing damaged vessels and contribute to the formation of new vessels.

Keywords: hypoxia • retinal neovascularization • growth factors/growth factor receptors