The behavior of endothelial cells on polyurethane nanocomposites and the associated signaling pathways

A series of nanocomposites from polyurethane (PU) incorporated with various low concentrations (17.4–174 ppm) of gold nanoparticles (approximately 5 nm) (denoted “PU–Au”) were used as a model system to study the mechanisms that influenced endothelial cell (EC) migration on biomaterial surfaces. The migration rate of ECs on the PU–Au nanocomposites was determined by a real-time image system. It was found that ECs had the highest migration rate on the nanocomposite containing 43.5 ppm of gold (“PU–Au 43.5 ppm”). The high EC migration rate was associated with increased levels of endothelial nitric oxide synthase (eNOS) and phosphorylated-Akt (p-Akt) expressed by ECs cultured on PU–Au. The inductions of both eNOS and p-Akt on PU–Au were abolished by the addition of LY294002 (PI3K inhibitor), suggesting that these cellular events may be regulated through the PI3K signaling pathway. Using a biotinylated VEGF-165 that recognizes VEGF receptors and by FACS analysis, slightly higher expression of VEGF receptors for ECs on PU–Au was also demonstrated. Phalloidin staining showed that actin appeared as a circumferential band surrounding each cell on tissue culture polystyrene, whereas on PU–Au, especially on PU–Au 43.5 ppm, the cells had their margin spread out and extend processes with stress fibers in the protruding lamellipodia. Moreover, the higher EC migration rate on PU–Au 43.5 ppm was suppressed by LY294002. The higher protein expression of focal adhesion kinase (FAK) on PU–Au 43.5 ppm was observed in FAK-GFP transfected ECs. It was concluded that PU–Au nanocomposites activated FAK and the PI3K/Akt signaling pathway in ECs, leading to proliferation and migration of ECs on these surfaces.