Abstract 561: Human Stem Cell--Derived Models of Endothelial Cell Morphogenesis: Label-Free Analysis of Invasion, Migration and Sprouting Angiogenesis

Abstract

Endothelial cell (EC) morphogenesis during angiogenesis is a highly dynamic process tightly regulated by growth factor activity in the extracellular milieu. Current in vitro models of EC morphogenesis employ endpoint analyses of primary endothelial cells, such as HUVECs. These vascular cells can exhibit functional variability between donors and methods for measuring morphogenesis in response to soluble cues typically employ potentially destructive labeling and thus fail to capture the full complexity of processes involved in EC morphogenesis. Here, we describe use of human induced pluripotent stem cell derived endothelial cells (iPSC-ECs) in a real-time electronic cell sensor impedance array technology to monitor invasion and migration behavior. Additionally, an image processing algorithm was developed to assess sprouting behavior from aggregated iPSC-EC organoid cultures. iPSC-EC responses on the impedance platform assessed the impact of serum, growth factors ( e.g. VEGF, FGF-2), and small-molecule angiogenesis inhibitors ( e.g. Sunitinib, SU5402) on the proliferation, migration, and invasion of iPSC- ECs compared to primary HUVECs. Our results indicated that iPSC-ECs were more sensitive to inhibition of VEGF mediated invasion by SU5402 than HUVECs with EC50s of 4.4nM and 99nM, respectively. In addition, matrigel invasion toward VEGF was not observed appreciably with HUVECs, whereas iPSC-ECs demonstrated a marked invasive phenotype that could be inhibited by Nocodazole at 1nM. Using a novel method to monitor the sprouting behavior of the iPSC-ECs in an aggregated organoid into matrigel by HCA and image processing, IC50s for SU5402 (2.7nM) and Nocodazole (0.4nM) were determined. The use of iPSC-derived endothelial cells provides a robust and reproducible source of cells that outperform HUVECs. The real-time monitoring of cellular processes offers important advantages over traditional end-point assays - specifically, the ability to measure receptor activation along with morphologic and adhesive remodeling of EC upon growth factor activation and inhibition. The combination of iPSC- ECs together with real time monitoring systems provides a more biologically relevant human model system for vasculogenesis investigations.