In vitro and In vivo imaging of antivasculogenesis induced by Noggin protein expression in human venous endothelial cells

Abstract

Noggin protein is a potent bone morphogenetic protein (BMP) antagonist capable of inhibiting vasculogenesis even in the presence of provasculogenic VEGF and FGF-2. We found that human umbilical vein endothelial cells (HUVECs) do not express Noggin in culture and used these cells for modeling of antivasculogenesis. We hypothesized that high-efficiency transduction of HUVECs with bicistronic lentiviral vector encoding Noggin and enhanced green fluorescent protein (EGFP) enables direct visualization of Noggin effects in homogenous primary cell populations in vitro and in vivo. By comparing HUVECs transduced with a control GFP and GFP/Noggin expression cassettes, we showed that constitutive and orthotopic Noggin protein expression did not influence cell proliferation, down-regulated BMP-2 expression, and showed no effect on BMP receptor transcripts. We demonstrated that in contrast to GFP-only control, Noggin expression in endothelial cells abrogated endothelial migration in response to monolayer injury, blocked endothelial transmigration, and caused abrogation of cord formation in vitro. Adding exogenous BMP-4 restored the formation of cords. Imaging experiments in vivo investigated vessel formation in Matrigel implants in athymic mice by utilizing GFP imaging or magnetic resonance imaging of perfusion in the implants. Both approaches demonstrated the lack of functional vessel formation after the adoptive transfer of GFP/Noggin-expressing human endothelial cells in mice.