Figure 7.
Fluorescent microscope images showing antiapoptotic effect of human angiopoietin-1 and human VEGF on HUVECs at day 14. Bar, 50 μm. It was observed that on day 14, the proteins coencapsulated in the nanoparticles significantly decreased HUVEC apoptosis compared with the control, as well as the individual proteins. Human vascular endothelial cells were seeded onto a 96-well plate at a cell density of 2 × 104 cells per well. The medium was then replaced with 0.1 mL of A) nanoparticle supernatant, B) nanoparticle supernatant with excess human VEGF antibody, C) nanoparticle supernatant with excess human angiopoietin-1 antibody and incubated for 96 hours. D) Control cultures received the same amount of serum-free media without addition of human VEGF and human angiopoietin-1 proteins. The numbers of cells floating in each well were collected after phosphate-buffered saline washing and counted. The number of apoptotic cells in the adherent cells was then determined after 96 hours using the MitoTracker® Red CMXRos kit and DAPI nucleic acid staining and counted using fluorescence microscopy.
Abbreviations: HUVECs, human vascular endothelial cells; VEGF vascular endothelial growth factor.