Nerve growth factor promotes endothelial progenitor cell-mediated angiogenic responses.

PURPOSE: In response to ischemia, retinal neuronal cells express nerve growth factor (NGF), which can be proangiogenic. Endothelial progenitor cells (EPCs) can participate with the resident vasculature to promote angiogenesis. We postulated that NGF may stimulate CD34⁺ EPCs to convert to an angiogenic phenotype. METHODS: Human CD34⁺ cells and human retinal endothelial cells (HRECs) were used to examine the effect of NGF on key steps associated with neovascularization. CD34⁺ cells and HRECs were stimulated with NGF (1 to 4 pM) for 24, 48, and 72 hours. Cell migration was measured using a modified Boyden chamber assay. Expression of the receptor for the cytokine stromal derived growth factor 1 (SDF-1), CXCR-4, was assessed by flow cytometry. In vitro angiogenesis was tested using a three-dimensional (3D) extracellular matrix with HRECs/CD34⁺ cell cocultures. NGF receptor activation was assessed by western analysis. RESULTS: NGF promoted proliferation of CD34⁺ cells but not HRECs. Pretreatment of CD34⁺ cells with NGF increased CXCR-4 expression in CD34⁺ cells, resulting in enhanced migration to SDF-1 (P < 0.0001). The enhanced tubule-forming effect of NGF in HRECs was further potentiated by coculture with NGF-pretreated CD34⁺ cells (P < 0.01). The beneficial effect of NGF was blocked (P < 0.0001) by the ERK inhibitor PD98059. In both CD34⁺ and HRECs, NGF increased phosphorylation of neurotrophic tyrosine kinase receptor type 1 (TrkA) receptor by ERK1 activation (P < 0.01). CONCLUSIONS: Our in vitro results suggest that NGF released from ischemic nerves in vivo may contribute to the "angiogenic switch" by stimulating the angiogenic behavior of CD34⁺ cells while minimally affecting resident retinal endothelial cells.