Figure 8

Increased VEGF expression in endothelial cells that form tubular structures. DD-1 cells were cultured in basal media (BM, without endothelial growth supplements), basal media under conditions of low calcium (0.1 mM EDTA), or basal media with EDTA and nicotine (10 μM), an activator of NFAT transcription factors, for 6 h. Cell-free supernatants were collected by centrifugation and cellular debris was removed by filtration. The levels of VEGF in the culture supernatants (sVEGF) were determined using a commercial ELISA following the manufacturer's instructions. VEGF gene expression was examined by Northern blotting. Ribosomal RNA bands (28S and 18S) stained with ethidium bromide are shown as loading controls.