IGFBP-3 Regulates Hematopoietic Stem Cells (HSC) and Endothelial Precursor Cell Differentiation, Migration and Retinal Development

Abstract

Purpose:: Circulating endothelial precursor cells (EPCs) derived from bone marrow hematopoietic stem cells (HSCs) home to areas of hypoxia and participate in vessel repair. Since most homing factors are expressed by hypoxic regions of , we asked whether local expression of the hypoxia regulated protein insulin-like growth factor binding protein-3 (IGFBP-3), could stimulate EPC homing and vascular repair functions.

Methods:: In vitro effects of IGFBP-3 on EPCs were tested using the Boyden chamber assay and tube formation on fibronectin. A plasmid expressing IGFBP-3 under the control of a proliferating endothelial cell-specific promoter was injected intravitreally into neonatal pups undergoing the oxygen-induced retinopathy model. Separate pups were also injected with green fluorescent protein (gfp)-expressing mouse HSCs transfected with the same IGFBP-3 expressing plasmid. Retinas were examined on post-natal day 17 using confocal microscopy.

Results:: Exposure of the CD34⁺ EPC population, to IGFBP-3 resulted in rapid differentiation into endothelial cells (confirmed by incorporation of fluorescently labeled acetylated LDL) and a dose-dependent increase in cell migration and capillary tube formation. Injection of IGFBP-3 plasmid alone or injection of HSC transfected with the plasmid resulted in protection of the developing vasculature from hyperoxic injury. Quantitative analysis of vascular density in this experiment supported our findings. One-way ANOVA showed that IGFBP-3 significantly protects the retinal vasculature from hyperoxia-induced vessel regression in mid peripheral [F (2, 42) = 36.98, P<0.001] and peripheral regions, but did not have any significant effect on the central region of the retina. Preretinal neovascularization was reduced by 70 ± 10% (P < 0.001) in pups injected with IGFBP-3 plasmid compared to controls. In normoxia mice increasing IGFBP-3 expression quickened the rate of retinal vascular development.

Conclusions:: These studies support that IGFBP-3 has profound effects on HSCs and EPCs promoting their differentiation into endothelial cells and their participation in vascular repair, representing a physiological adaptation to ischemia and potentially a novel therapeutic target for treatment of ischemic conditions.

Keywords: hypoxia • retinal neovascularization • differentiation